In vitro EVALUATION OF EUCALYPTUS ECTOMYCORRHIZAE ON SUBSTRATE WITH PHOSPHORUS DOSES FOR FUNGAL PRE-SELECTION

The benefit promoted by ectomycorrhizal depends on the interaction between symbionts and phosphorus (P) contents. Phosphorus effect on ectomycorrhizal formation and the effectiveness of these in promoting plant growth for fungal pre-selection were assessed under in vitro conditions. For P effect evaluation, Eucalyptus urophylla seedlings inoculated with four Pisolithus sp. isolates and others non-inoculated were grown on substrate containing 0.87, 1.16 and 1.72 mg P per plant. For evaluation of effectiveness and fungal pre-selection, other 30 isolates of Pisolithus sp., Pisolithus microcarpus ITA06 isolate, Amanita muscaria AM16 isolate, Scleroderma areolatum SC129 isolate were studied. D26 isolate promoted the highest plant heights for the three P doses, D51 at the lower dose and D72 at the intermediate dose. P doses did not influenced shoot fresh weight and fungal colonization. In the pre-selection of fungi, 14 isolates of Pisolithus sp., P. microcarpus ITA06 isolate and S. areolatum SC129isolate increased plant height and fresh weight. D82 isolate of Pisolithus sp. had effect singly on plant height while D17 and D58 on fresh weight. Of these, only D15, D17, D58 and ITA06 had typical ectomycorrhizae. The cultivation in vitro has shown adequate for pre-selection of ectomycorrhizal fungi. Colonization and benefits depend on species and isolate. D15, D17 and D58 of Pisolithus sp. and P. microcarpus isolate ITA06 are the most promising for nursery studies.

TEMPERATURE AND SUBSTRATE INFLUENCES ON SEED GERMINATION AND SEEDLING FORMATION IN Callisthene fasciculata Mart. (VOCHYSIACEAE) IN THE LABORATORY1

ABSTRACTCallisthene fasciculata Mart. is a tree belonging to the Vochysiaceae family. Its wood is moderately heavy and resistant and used to make poles, beams, and other structures. The aim of this work was to evaluate seed germination and the initial growth of seedlings of C. fasciculata at different temperatures and in different substrates. Seeds were collected from fruits in the Pantanal de Miranda, Mato Grosso do Sul state, Brazil. In one experiment, the seeds were subjected to constant temperatures of 20, 25, 30 and 35 °C and to alternating temperatures of 20-30 and 25-35 °C (on paper substrate). In another experiment, the seeds were subjected to temperatures of 20 and 25 °C on three substrates (sand, vermiculite and between paper) in a germinator. The experiment had a randomized design, with four replicates of 25 seeds per treatment. The F-values obtained for germination indicated no significant effect of substrate or temperature on the final germination percentage. The analyses revealed no effect of a treatment interaction (temperature x substrate) on either germination or average germination time; however, a treatment interaction effect was observed on the germination speed index. The treatment combinations yielding the best performance were between paper substrate at 20 °C and sand substrate at 25 °C. There was a significant effect of the interaction between temperature and substrate on seedling growth, with increased root growth observed in the between paper substrate at 25 °C and increased aerial component growth in both sand at 20 °C and vermiculite at 25 °C. The between paper treatment at 25 °C yielded the greatest final seedling size. Between paper is the most recommended substrate for the production of seedlings due to its ease of handling and lower probability of contamination.

ROOTING MINI-CUTTINGS OF Paulownia fortunei var. mikado DERIVED FROM CLONAL MINI-GARDEN1

ABSTRACTWe aimed to evaluate the technical efficiency of mini-cuttings technique on vegetative propagation of Paulownia fortunei (Seem.) Hemsl. var. Mikado, as well as the possible existence of anatomical barriers to its rooting. Therefore, plants originated from cuttings formed the mini-stumps and, consequently the clonal mini-garden, which was conducted in semi-hydroponic system. We evaluated the survival of mini-stumps and sprouts production for five successive collects, the percentage of mini-cuttings rooting and their anatomical description. The mini-cuttings were prepared with about 6 to 8 cm in length and two leaves reduced by about 50% in the upper third, being remained an area of, approximately 78 cm2 (10 cm diameter). The mini-cuttings were placed in tubes of 53 cm3, with substrate formed with fine vermiculite and carbonized rice hulls (1:1 v/v) and rooted in acclimatized greenhouse. After 30 days we evaluated the percentage of rooted mini-cuttings, radicial vigor (number and length of roots / mini-cutting), callus formation, emission of new shoots and maintenance of the original leaves. The mini-stumps showed 100% survival after five collects and an average production of 76-114 mini-cuttings/m2/month and rooting ranged from 70 to 90%. Mini-cuttings technique is efficient in to propagate adult propagules of the species and there are not anatomical barriers preventing roots emission.

SUBSTRATE, LIME, PHOSPHORUS AND TOPDRESS FERTILIZATION IN MACAW PALM SEEDLING PRODUCTION

ABSTRACT The macaw palm [Acrocomia aculeata (Jacq.) Lood. ex Mart] has been domesticated to subsidize biodiesel production programs in Brazil. However, little is known about the seedling production of this species. This study aimed to evaluate substrate mixtures, limestone and phosphorus rates for substrate amendment and topdressing frequency in macaw palm seedlings. Three trials were conducted in a greenhouse up to six months of nursery cultivation. Trial 1: determination of percent mineral and organic fractions of seven substrate mixtures. Trial 2: evaluation of four limerates for soil amendment versus four phosphorus rates. Trial 3: evaluation of N, K and Mg topdressing frequency. Significant differences were found in the three trials for most of the variables (plant height, leaf number, shoot dry mass, root dry mass, vigor and bulb diameter). The main results obtained were as follow: Trial1 - the best seedling growth was observed in substrates with at least 25% organic matter. Trial2 -lime rates ranging from 0.50 to 1.25 kg associated with 3 to 4 kg of single superphosphate per m3 of substrate provided the best seedling growth. Trial 3 - topdressing fertilization provided better development of seedlings regardless of frequency.

Substrate with Organosuper® for cucumber seedlings formation in protected environments and polystyrene trays

Culture environments, trays and doses of organic compost were evaluated in the formation of cucumber seedlings (Cucumis sativus L.). Five environmental conditions were tested: (A1) a greenhouse with height of 2.5 m, covered with polyethylene film, (A2) nursery with height of 2.5 m, monofilament fabric, 50% shading, (A3) nursery with height of 2.5 m, heat-reflective screen, 50% shading, (A4) nursery with a height of 1.8 m, covered with coconut tree straw and (A5) greenhouse with height of 4.0 m, covered with polyethylene film, with zenith opening and thermo-reflective cloth under the plastic. Trays of 72 (R1) and 128 (R2) cells were filled with 93% soil and 7% organic compound (S1), 86% soil and 14% organic compound (S2) and 79% soil and 21% organic compound (S3). It was used a randomized design in split-split-plot scheme, with five replicates (environments x trays x substrates). The greenhouses provide the best environments for the formation of cucumber seedlings. A tray of 72 cells is the best container, promoting more vigorous seedlings in substrate with soil and 7 or 14% organic compound.

Anaerobic reactors with biofilter and different diameter-length ratios in cassava starch industry wastewater treatment

The use of anaerobic reactors with media support in the treatment of wastewater from the cassava starch industry has emerged as a viable option because it allows the application of high organic loads and a significant reduction of the HDT needed for the treatment. This research aimed at studying the process of biodigestion in two anaerobic reactors with bamboo support, in the treatment of effluent of cassava starch, by evaluating their performance. The two reactors used present the following diameter: length ratio, 1:6 and 1:3. The organic loads applied to the systems were 0.519, 1.156, 1.471, 3.049, 4.347, 4.708 and 5.601g.L-1.d-1. Regarding the efficiency of removal of COD, TS and TVS, no statistically significant differences were obtained between the reactors. The two systems evaluated showed a stable behavior with respect to the VA/TA (volatile acidity/total alkalinity) for all submitted loads. The reactors tended to the maintenance of biogas production as a function of consumed COD for the last three organic loads applied, indicating an ability to withstand higher organic loads.

Spatial distribution and preferential substrate of Neoregelia johannis (Carriére) L.B. Smith (Bromeliaceae) in a disturbed area of Atlantic Rainforest at Ilha Grande, RJ, Brazil

In bromeliad populations, amount of light and available substrates influence individuals spatial organization. In Atlantic Rainforest of Ilha Grande, the heliophylous bromeliad Neoregelia johannis is a large and abundant species. In this forest, it would be expected that N. johannis would occupy stable substrates, as large trunks, large branches, rock boulders or ground, with high sunlight, enough for the bromeliad survivor. In the present work, we analyzed the distribution and most used substrates of N. johannis in secondary forest. We analyzed the frequency of reproductive modes (sexual and vegetative) used by the bromeliad shoots, registering if the shoots were originated from seeds or by vegetative reproduction. The results indicated an aggregated distribution pattern (Ip = 0.052). The preferred substrate was boulders (91%), whereas tree trunks (6%) and the ground (3%) were rarely used. Small and fragile substrates are unstable to support large adults of this species, which may explain the predominant pattern of establishment over boulders within the secondary forest, as the presence of this substrate also results in more opened canopy cover. Approximately 50% of young individuals entered the population by vegetative reproduction. We conclude that the preferential habit and the aggregated distribution of N. johannis are due to the conjunction of preferred substrate with higher amount of light resulting from breaks in tree canopy over areas with rock blocks, and high frequency of recruitment by vegetative reproduction.

Synchrony is more than overlap: measuring phenological synchronization considering time length and intensity

The degree of flowering and fruiting synchronization is believed to have ecological and evolutionary relevance at several scales. Here we discuss some measures that have been used to estimate synchrony and propose an index that incorporates both the entire length of an individual phenophase and variation in the number of flowers or fruits over that time period. This new index describes more accurately the phenological synchrony among individuals and populations.

Family names and the length of the Y chromosome in Brazilian blacks

The variability of the lengths of the heterochromatic and euchromatic segments of the human Y chromosome was studied by a quantitative method of densitometric measurement in 60 normal and unrelated black individuals (30 with and 30 without devotional surnames), living in Salvador, Bahia, northeastern Brazil. Thirty normal and unrelated Caucasian individuals of European origin, living in Curitiba, Paraná, south Brazil, were included as controls. The heterochromatic segment and total Y chromosome lengths were greater in caucasians than in blacks without devotional surnames, and these were greater than in blacks with devotional surnames. These findings are in agreement with previous reports of a higher percentage of black ancestry in blacks carrying devotional surnames than those carrying non-devotional ones.

Construction of occluded recombinant baculoviruses containing the full-length cry1Ab and cry1Ac genes from Bacillus thuringiensis

The administration of baculoviruses to insects for bioassay purposes is carried out, in most cases, by contamination of food surfaces with a known amount of occlusion bodies (OBs). Since per os infection is the natural route of infection, occluded recombinant viruses containing crystal protein genes (cry1Ab and cry1Ac) from Bacillus thuringiensis were constructed for comparison with the baculovirus prototype Autographa californica nucleopolyhedrovirus (AcNPV). The transfer vector pAcUW2B was used for construction of occluded recombinant viruses. The transfer vector containing the crystal protein genes was cotransfected with linearized DNA from a non-occluded recombinant virus. The isolation of recombinant viruses was greatly facilitated by the reduction of background "wild type" virus and the increased proportion of recombinant viruses. Since the recombinant viruses containing full-length and truncated forms of the crystal protein genes did not seem to improve the pathogenicity of the recombinant viruses when compared with the wild type AcNPV, and in order to compare expression levels of the full-length crystal proteins produced by non-occluded and occluded recombinant viruses the full-length cry1Ab and cry1Ac genes were chosen for construction of occluded recombinant viruses. The recombinant viruses containing full-length and truncated forms of the crystal protein genes did not seem to improve its pathogenicity but the size of the larvae infected with the recombinant viruses was significantly smaller than that of larvae infected with the wild type virus.

The insulin receptor substrate 1 associates with phosphotyrosine phosphatase SHPTP2 in liver and muscle of rats

Insulin stimulates the tyrosine kinase activity of its receptor resulting in the phosphorylation of its cytosolic substrate, insulin receptor substrate-1 (IRS-1) which, in turn, associates with proteins containing SH2 domains. It has been shown that IRS-1 associates with the tyrosine phosphatase SHPTP2 in cell cultures. While the effect of the IRS-1/SHPTP2 association on insulin signal transduction is not completely known, this association may dephosphorylate IRS-1 and may play a critical role in the mitogenic actions of insulin. However, there is no physiological demonstration of this pathway of insulin action in animal tissues. In the present study we investigated the ability of insulin to induce association between IRS-1 and SHPTP2 in liver and muscle of intact rats, by co-immunoprecipitation with anti-IRS-1 antibody and anti-SHPTP2 antibody. In both tissues there was an increase in IRS-1 association with SHPTP2 after insulin stimulation. This association occurred when IRS-1 had the highest level of tyrosine phosphorylation and the decrease in this association was more rapid than the decrease in IRS-1 phosphorylation levels. The data provide evidence against the participation of SHPTP2 in IRS-1 dephosphorylation in rat tissues, and suggest that the insulin signal transduction pathway in rat tissues is related mainly to the mitogenic effects of the hormone.

Erythrocyte glucose-6-phosphate dehydrogenase activity assay and affinity for its substrate under "physiological" conditions

Glucose-6-phosphate dehydrogenase (G6PD) activity and the affinity for its substrate glucose-6-phosphate were investigated under conditions similar to the physiological environment in terms of ionic strength (I: 0.188), cation concentration, pH 7.34, and temperature (37oC). A 12.4, 10.4 and 21.4% decrease was observed in G6PD B, G6PD A+ and G6PD A- activities, respectively. A Km increase of 95.1, 94.4 and 95.4% was observed in G6PD B, G6PD A+ and G6PD A-, respectively, leading to a marked decrease in affinity. In conclusion, the observation of the reduced activity and affinity for its natural substrate reflects the actual pentose pathway rate. It also suggests a much lower NADPH generation, which is crucial mostly in G6PD-deficient individuals, whose NADPH availability is poor.

Characterization of six rat strains (Rattus norvegicus) by mitochondrial DNA restriction fragment length polymorphism

Restriction fragment length polymorphism (RFLP) was used to examine the extent of mtDNA polymorphism among six strains of rats (Rattus norvegicus) - Wistar, Wistar Munich, Brown Norway, Wistar Kyoto, SHR and SHR-SP. A survey of 26 restriction enzymes has revealed a low level of genetic divergence among strains. The sites of cleavage by EcoRI, NcoI and XmnI were shown to be polymorphic. The use of these three enzymes allows the 6 strains to be classified into 4 haplotypes and identifies specific markers for each one. The percentage of sequence divergence among all pairs of haplotypes ranged from 0.035 to 0.33%, which is the result of a severe population constriction undergone by the strains. These haplotypes are easily demonstrable and therefore RFLP analysis can be employed for genetic monitoring of rats within animal facilities or among different laboratories.

P-glycoprotein-mediated resistance to chemotherapy in cancer cells: using recombinant cytosolic domains to establish structure-function relationships

Resistance to chemotherapy in cancer cells is mainly mediated by overexpression of P-glycoprotein (Pgp), a plasma membrane ATP-binding cassette (ABC) transporter which extrudes cytotoxic drugs at the expense of ATP hydrolysis. Pgp consists of two homologous halves each containing a transmembrane domain and a cytosolic nucleotide-binding domain (NBD) which contains two consensus Walker motifs, A and B, involved in ATP binding and hydrolysis. The protein also contains an S signature characteristic of ABC transporters. The molecular mechanism of Pgp-mediated drug transport is not known. Since the transporter has an extraordinarily broad substrate specificity, its cellular function has been described as a "hydrophobic vacuum cleaner". The limited knowledge about the mechanism of Pgp, partly due to the lack of a high-resolution structure, is well reflected in the failure to efficiently inhibit its activity in cancer cells and thus to reverse multidrug resistance (MDR). In contrast to the difficulties encountered when studying the full-length Pgp, the recombinant NBDs can be obtained in large amounts as soluble proteins. The biochemical and biophysical characterization of recombinant NBDs is shown here to provide a suitable alternative route to establish structure-function relationships. NBDs were shown to bind ATP and analogues as well as potent modulators of MDR, such as hydrophobic steroids, at a region close to the ATP site. Interestingly, flavonoids also bind to NBDs with high affinity. Their binding site partly overlaps both the ATP-binding site and the steroid-interacting region. Therefore flavonoids constitute a new promising class of bifunctional modulators of Pgp.

Effect of practice and span length on the dual-task coordination executive test

The measure "mu", proposed as an index of the ability to coordinate concurrent box-crossing (BC) and digit-span (DS) tasks in the dual task (DT), should reflect the capacity of the executive component of the working memory system. We investigated the effect of practice in BC and of a change in the digit span on mu by adding previous practice trials in BC and diminishing, maintaining or increasing the digit sequence length. The mu behavior was evaluated throughout three trials of the test. Reported strategies in digit tasks were also analyzed. Subjects with diminished span showed the best performance in DT due to a stable performance in DS and BC in the single- and dual-task conditions. These subjects also showed a more stable performance throughout trials. Subjects with diminished span tended to employ effortless strategies, whereas subjects with increased span employed effort-requiring strategies and showed the lowest means of mu. Subjects with initial practice trials showed the best performance in BC and the most differentiated performance between the single- and dual-task conditions in BC. The correlation coefficient between the mu values obtained in the first and second trials was 0.814 for subjects with diminished span and practice trials in BC. It seems that the within-session practice in BC and the performance variability in DS affect the reliability of the index mu. To control these factors we propose the introduction of previous practice trials in BC and a modification of the current method to determine the digit sequence length. This proposal should contribute to the development of a more reliable method to evaluate the executive capacity of coordination in the dual-task paradigm.